title: Identification of HIV-1 reverse transcriptase inhibitors using a scintillation proximity assay.
abstract: The human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) converts the viral single-stranded RNA into double-stranded DNA. The inhibition of reverse transcription in the viral life cycle has proven its efficacy as a clinically relevant antiviral target, but the appearance of resistance mutations remains a major cause of treatment failure and stresses the continuous need for new antiviral compounds. In this chapter, we describe an HIV-1 RT scintillation proximity assay (SPA) to identify inhibitors of the RT. The assay uses an RNA/DNA (poly(rA)/oligo(dT)) template/primer bound to SPA beads, which contain scintillant. Reverse transcriptase extends the primer by incorporating [(3)H]dTTP and dTTP, which results in light production by the scintillant in the bead. Compounds that inhibit reverse transcriptase will prevent the incorporation of tritiated dTTP resulting in a reduction of emitted light compared to the untreated controls.
authors.affiliation: Janssen Infectious Diseases-Diagnostics BVBA, Johnson & Johnson Corporation, Beerse, Belgium.
authors.forename: Bertrand
authors.initials: B
authors.lastname: Van Schoubroeck
authors.forename: Marnix
authors.initials: M
authors.lastname: Van Loock
authors.forename: Tania
authors.initials: T
authors.lastname: Ivens
authors.forename: Pascale
authors.initials: P
authors.lastname: Dehertogh
authors.forename: Dirk
authors.initials: D
authors.lastname: Jochmans
authors.forename: Géry
authors.initials: G
authors.lastname: Dams
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors*; D015497:HIV-1 / Q000187:drug effects* / Q000201:enzymology*; D006801:Humans; D020128:Inhibitory Concentration 50; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D012588:Scintillation Counting / Q000379:methods*
chemical_list: D019380:Anti-HIV Agents; D018894:Reverse Transcriptase Inhibitors; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase